The G protein-coupled prostaglandin F2α (PGF2α) receptor [F prostanoid (FP) receptor] has been implicated in many physiological events including cardiovascular, respiratory, immune, reproductive, and endocrine responses. Binding of PGF2α to FP receptor elicits inositol production and protein kinase C-dependent MAPK activation through Gα(q) coupling. Here we report that AL-8810, previously characterized as an orthosteric antagonist of PGF2α-dependent, Gα(q)-mediated signaling, potently activates ERK1/2 in a protein kinase C-independent manner. Rather, AL-8810 promoted ERK1/2 activation via an epidermal growth factor receptor transactivation mechanism in both human embryonic kidney 293 cells and in the MG-63 osteoblast-like cells, which express endogenous FP receptors. Neither AL-8810- nor PGF2α-mediated stimulation of FP receptor promoted association with β-arrestins, suggesting that MAPK activation induced by these ligands is independent of β-arrestin's signaling scaffold functions. Interestingly, the spatiotemporal activation of ERK1/2 promoted by AL-8810 and PGF2α showed almost completely opposite responses in the nucleus and the cytosol. Finally, using [(3)H]thymidine incorporation, we noted differential regulation of PGF2α- and AL-8810-induced cell proliferation in MG-63 cells. This study reveals, for the first time, the signaling biased nature of FP receptor orthosteric ligands toward MAPK signaling. Our findings on the specific patterns of ERK1/2 activation promoted by FP receptor ligands may help dissect the distinct roles of MAPK in FP receptor-dependent physiological responses.